Transition metal (II) complexes of hydrazones derived from tetralone: synthesis, spectral characterization, in vitro antimicrobial and cytotoxic studies

Article abstract of DOI:10.1007/s11164-021-04413-x

Abstract: A series of transition metal (II) complexes with general formula [M(L^1?4)₂(H₂O)₂] (where M = Co(II), Ni(II), Cu(II) and Zn(II)) was synthesized by the reaction of metal(II) acetates with hydrazones [HL¹–HL⁴] obtained from condensation of tetralone with hydrazide derivatives. The characterization of synthesized compounds (1–20) was done by using elemental analysis, different spectral studies (UV–Vis, ¹H NMR, ¹³C NMR, FT-IR, mass, ESR and fluorescence), magnetic susceptibility, molar conductance measurement and thermal (DTA, TGA and DTG) analysis. The characterization results suggested the bidentate nature of the hydrazones, which chelate with the metal ion via nitrogen of (C=N) group and deprotonated carbonyl oxygen in the enolized form, resulting in stable, non-volatile octahedral complexes. The antimicrobial potential of the compounds was evaluated against four bacterial, i.e. (S. aureus, B. subtilis, P. aeruginosa and E. coli), and two fungal species, i.e. (A. niger and C. albicans), by a serial dilution method with complexes 15 and 16 as most active compounds against microbes. All the compounds were also tested for its cytotoxicity (in vitro) against three human cancer cell lines (A549, HeLa and MCF-7) and one normal cell line (L₆) by the MTT assay, which focussed on an increased activity of compounds 7 and 15 towards A549, MCF-7 and HeLa cancer cell line with IC₅₀ values ranging from 10.76 to 16.42?μg/mL, and compounds were found to be non-toxic towards L₆ cell line as compared to the standard drug doxorubicin. The results demonstrated that complexation enhanced the biological activity of compounds. Graphic abstract: The synthesized compounds (1–20) were screened for antitumor activities against A549, MCF7, Hela cancer cell lines. Copper complex (7) and (15) was found to be the most active antitumor agent and less toxic against L6—Rat myoblast normal cell line than standard doxorubicin.[Figure not available: see fulltext.].

Full text of DOI:10.1007/s11164-021-04413-x

Research on Chemical Intermediates
https://doi.org/10.1007/s11164-021-04413-x
Transition metal (II) complexes of hydrazones derived
from tetralone: synthesis, spectral characterization, in vitro
antimicrobial and cytotoxic studies
J. Devi1 · S. Sharma1,2 · S. Kumar1 · D. K. Jindal3 · P. P. Dutta4,6 · D. Kumar5
Received: 14 November 2020 / Accepted: 30 January 2021
© Springer Nature B.V. 2021
Abstract
A series of transition metal (II) complexes with general formula [M(L1−4)2(H2O)2]
(where M=Co(II), Ni(II), Cu(II) and Zn(II)) was synthesized by the reaction of
metal(II) acetates with hydrazones [HL1–HL4] obtained from condensation of
tetralone with hydrazide derivatives. The characterization of synthesized compounds
(1–20) was done by using elemental analysis, diferent spectral studies (UV–Vis,
1H NMR, 13C NMR, FT-IR, mass, ESR and fuorescence), magnetic susceptibility,
* J. Devi
jaya.gju@gmail.com
S. Sharma
somsamainia@gmail.com
S. Kumar
sanjeevbagri2221@gmail.com
D. K. Jindal
deepakjindal44@yahoo.com
P. P. Dutta
partha184@gmail.com
D. Kumar
guptadeepak002@gmail.com
1
Department of Chemistry, Guru Jambheshwar University of Science and Technology,
Hisar 125001, India
2
Department of Chemistry, Government College Hisar, Hisar 125001, India
3
Department of Pharmaceutical Sciences, Guru Jambheshwar University of Science
and Technology, Hisar 125001, India
4
Institute of Advanced Studies in Science & Technology, Life Science Division,
Guwahati 781035, Assam, India
5
School of Pharmaceutical Sciences, Shoolini University, Solan 173212, Himachal Pradesh,
India
6
Faculty of Pharmaceutical Science, Assam Down Town University, Panni khaiti,
Guwahati 781026, Assam, India
Vol.:(0123456789)
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J. Devi et al.
molar conductance measurement and thermal (DTA, TGA and DTG) analysis. The
characterization results suggested the bidentate nature of the hydrazones, which
chelate with the metal ion via nitrogen of (C=N) group and deprotonated carbonyl
oxygen in the enolized form, resulting in stable, non-volatile octahedral complexes.
The antimicrobial potential of the compounds was evaluated against four bacterial,
i.e. (S. aureus, B. subtilis, P. aeruginosa and E. coli), and two fungal species, i.e.
(A. niger and C. albicans), by a serial dilution method with complexes 15 and 16 as
most active compounds against microbes. All the compounds were also tested for its
cytotoxicity (in vitro) against three human cancer cell lines (A549, HeLa and MCF-
7) and one normal cell line (L6) by the MTT assay, which focussed on an increased
activity of compounds 7 and 15 towards A549, MCF-7 and HeLa cancer cell line
with IC50 values ranging from 10.76 to 16.42 μg/mL, and compounds were found
to be non-toxic towards L6 cell line as compared to the standard drug doxorubicin.
The results demonstrated that complexation enhanced the biological activity of
compounds.
Graphic abstract
The synthesized compounds (1–20) were screened for antitumor activities against
A549, MCF7, Hela cancer cell lines. Copper complex (7) and (15) was found to be
the most active antitumor agent and less toxic against L6—Rat myoblast normal cell
line than standard doxorubicin.
Keywords Hydrazone · Bidentate · Octahedral · Antimicrobial · Anticancer
1 3

Transition metal (II) complexes of hydrazones derived from…
Abbreviations
DME
M Dulbecco’s modifed Eagle’s medium
DMF
Dimethylformamide
DMSO
Dimethyl sulfoxide
DMSO-d6 Deuterated dimethyl sulfoxide
ESR
TGA
DTA
MS
Electron spin resonance
Thermal gravimetric analysis
Diferential thermal analysis
Mass spectra
FAB
FT-IR
A549
HeLa
MCF7
L6
Fast atom bombardment
Fourier transform infrared spectroscopy
Human lung carcinoma cell line
Human cervical carcinoma cell line
Human breast carcinoma cell line
Rat myoblast cell line normal
MTT
NMR
IC50
3-(4,5-Dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide
Nuclear magnetic resonance
Concentration of drug required to inhibit the growth of 50% of tumour
cells
Introduction
Cancer or malignant neoplasm is one of the most widespread and highly feared
diseases for human life causing death nowadays in both developing and devel-
oped countries. Cancer results from the uncontrolled multiplication of ingenious
modifed normal human cells. Researchers throughout the globe have contributed
signifcantly towards the anticancer drug development research. Cisplatin (cis-
diamminedichloroplatinum(II)), a platinum metal-based drug, was a major develop-
ment in cancer treatment [1], but its adverse efects such as sickness, kidney and
liver failure, heavy metal toxicity and drug resistance [2] promoted researchers
to discover new metallodrugs that are less toxic, economic, target specifc with a
higher efciency.
In the last few decades, the applications of transition metal complexes for the
medical therapy of cancer are extensively investigated, and consequently, they were
acknowledged as promising anticancer agents. Transition metal complexes demon-
strated various therapeutic properties and are widely used in medicine as well as
have other applications also [3]. The ligand scafolds play an important role in the
chemotherapy, and among ligand system, hydrazones consisting of a carbon–nitro-
gen double bond (R–CO–NH–N=CHR) group [4] belong to special class due to
their interesting multidentate chelating properties and form complexes with almost
all the metal ions in diferent oxidation states depending upon the carbonyl group
and hydrazide derivatives [5, 6]. They possess diverse biological and pharmaceuti-
cal activities [7, 8] such as antimicrobial [9, 10], anticancer [11, 12] and antioxi-
dant properties [13]. They also show applications in wastewater treatment [14],
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J. Devi et al.
catalytical activity [15, 16], good magnetic behaviour [17], specifc sensors, non-
linear optical devices [18], resin for separation [19] and acting as enzyme inhibitor
[20].
Tetralone, carbonyl entity is selected here for hydrazones’ formation, represents
an important compound in modern medicinal chemistry as it has considerable
importance in biological feld [21], anti-trypanosoma, antidepressant in sertraline
drug, antimalarial [22], anticancer, antioxidant [23], anti-infammatory agent [24]
and as inhibitors of monoamine oxidase [25]. As a result to develop ingenious anti-
cancer agent, huge number of transition metal complexes were synthesized from
hydrazones and tested for anticancer activities, and many of them exhibited excel-
lent activities [26]. The literature survey suggested that biological potential of com-
pounds gets infuenced by the coordination number of metal atom, structure of mol-
ecules and substituents attached to metal atom.
Due to the widespread therapeutic and diagnostic properties of hydrazones
and tetralone derivatives, it seemed appealing to investigate compounds that have
remarkable applications of both antimicrobial and anticancer activities, so here we
have reported the synthesis of hydrazones from condensation of hydrazide deriva-
tives with tetralone and reacted them with transition metal(II) acetates to form
complexes. For characterization and structure determination, various spectroscopic
analyses (UV–Vis, 1H NMR, 13C NMR, FT-IR, mass, ESR and fuorescence), mag-
netic susceptibility, elemental analysis, molar conductance measurements and ther-
mal (TGA, DTG and DTA) methods were used. Screening of compounds for cyto-
toxic activity against three cancer cell lines—human breast adenocarcinoma cell line
(MCF7), human alveolar adenocarcinoma epithelial cell line (A549), human cervi-
cal cell line (HeLa) and Rat myoblast cell line normal (L6)—were assayed. The anti-
microbial activity was also tested against four bacterial, i.e. (S. aureus, B. subtilis, E.
coli and P. aeruginosa) and two fungal, i.e. (A. niger, C. albicans), strains.
Experimental
Materials
The solvents, chemicals and reagents used for the research were of analytical grade.
Hydrazide derivatives (nicotinic acid, benzoic acid, toluic acid and isonicotinic acid),
α-tetralone and metal(II) acetates [Co(CH3COO)2·4H2O, Ni(CH3COO)2·4H2O,
Cu(CH3COO)2·H2O and Zn(CH3COO)2·2H2O] were purchased from Sigma-Aldrich
company. The solvents used were dried as reported in the literature [27].
Instrumentation
Infrared spectra of hydrazones and their metal complexes were measured by using
KBr pellets on Shimadzu IR afnity-I 8000 FT-IR spectrophotometer within
the range of 400–4000 cm−1. NMR spectra (1H and 13C) of hydrazones and their
zinc(II) complexes were obtained in CDCl3 by taking TMS as an internal reference
1 3

Transition metal (II) complexes of hydrazones derived from…
on Bruker Avance II 400 MHz NMR spectrometer. The coupling constant (J) and
chemical shift (δ) were measured in Hz and ppm. Elemental (CHN) data were ana-
lysed by the instrument PerkinElmer 2400 elemental analyser. Standard gravimetric
procedures were used to determine the metal percentage in the compounds, nickel
as nickel dimethylglyoximate, zinc as zinc ammonium phosphate, cobalt as cobalt
pyridine thiocyanate and copper as cupreous thiocyanate [28]. Electronic spectral
data were obtained in DMSO by using UV–Vis–NIR Varian Cary-5000 spectro-
photometer. ESR spectral analysis of the Cu(II) complexes was measured by tak-
ing tetracyanoethylene (TCNE) free radical as an internal reference on the Varian
E112 X-band spectrometer. Gouy’s procedure is used to calculate the magnetic
susceptibilities of the compounds at 298 K by taking Hg[Co(SCN)4] as the refer-
ence to calibrate the instrument. Conductivity of 10−3 M solution was measured by
using a model-306 Systronics conductivity bridge at 298 K in DMSO. The melting
points (°C) of compounds were measured on a hot-stage Gallen Kamp melting point
apparatus in open capillaries. Mass spectra of compounds were obtained by using
API 2000 Applied Bio systems mass spectrometer in DMSO solvent. PerkinElmer
Diamond TG/DTA thermogravimetric analyser is used to carry out thermal (TGA,
DTG and DTA) study of compounds under nitrogen atmosphere with fow rate of
20 mL/min and heating rate 10 °C/min. Steady-state fuorescence spectra of hydra-
zones and their respective complexes were carried out on Shimadzu RF-5301 PC
spectrophotometer.
Pharmacology
Antimicrobial assay
Hydrazones and their respective transition metal complexes were screened for
their in vitro antimicrobial activity against four bacterial species: Bacillus subtilis
(MTCC 8561), Staphylococcus aureus (MTCC 2901), Pseudomonas aeruginosa
(MTCC 424), Escherichia coli (MTCC 16,521) and two fungal species: Candida
albicans (MTCC 227), Aspergillus niger (MTCC 8189) by serial dilution method
[29]. Test organisms were grown on nutrient agar (bacterial strain) and sabouraud
dextrose (fungal strain) as nutrient medium. The stock solution was obtained by add-
ing 2 mg of synthesized compounds in 10 mL DMSO. Further, to prepare a solu-
tion of 100 μg/mL concentration, 1 mL of this stock solution was poured into each
test tube having 1 mL of nutrient broth/sabouraud dextrose broth. The solutions of
50, 25, 12.50 and 6.25 μg/mL concentrations were prepared by the serial dilution
method. Microbial species were grown at 37 °C for 24 h in case of the bacterial and
at 25 °C for one week in case of the fungal strains. Ciprofoxacin is used as a refer-
ence drug for antibacterial and fuconazole for antifungal activity, and their activity
was also performed by taking DMSO as negative control. After incubation, the data
were observed at various concentrations and calculated their MIC values in μ mol/
mL. The microbial activity of compounds was carried out in duplicate to minimize
error, and an average value is reported here.
1 3

J. Devi et al.
Cytotoxic assay
The in vitro anticancer activity of the compounds (1–20) was tested as given by
Mosmann [30]. The cytotoxicity was evaluated against a series of three diferent
human carcinoma cell lines: A549 derived from human alveolar adenocarcinoma
epithelial cell lines (ATCC No.CCL-185), HeLa derived from human cervical ade-
nocarcinoma epithelial cell lines (ATCC No. CCL-2), MCF7 derived from human
breast adenocarcinoma epithelial cell lines (ATCC No.HTB-22) as well as normal
cell line L6 derived from rat skeletal muscle myoblast cells (ATCC No. CRL-1458),
and these cells were cultured in Dulbecco’s modifed Eagle’s medium (DMEM) sup-
plemented with 1% gentamicin, 10% foetal bovine serum (FBS) and 10% Pen-Strep.
Briefy, cells (1×106 per mL) were seeded in tissue culture grade multiwall plates
in complete medium and incubated under standard conditions in 37 °C humidifed
5% CO2 atmosphere. The complete medium was replaced after one day with FBS-
free medium and incubated overnight. The cells were then treated with the samples
in various concentrations into each well and incubated for 24 h. Well containing
medium alone (untreated cells) was served as a control. After the incubation period,
10 µL of MTT (5 mg/mL) was poured to each well, mixed thoroughly and incubated
again for 4 h. After the incubation period, cells were seen with the help of inverted
microscope for the existence of dark purple crystals of formazan at the bottom of the
wells. Isopropanol (100 µL) with 0.04 M HCl was poured to each well and mixed it
completely by repeated pipetting with a multi-channel pipette. The HCl converts the
phenol red in tissue culture medium to a yellow colour that does not interfere with
the MTT formazan measurement. The isopropanol dissolves the formazan to give a
homogeneous blue solution suitable for absorbance calculation, and absorbance was
calculated by ELISA plate reader (Filter Max F3 Multi-Mode Micro plate Readers,
Molecular Devices) with a test wavelength of 570 nm and a reference wavelength of
630 nm. The efect of the samples on the proliferation of cells was expressed as the
% inhibition of cell proliferation and measured as (Absorbance of control cells—
Absorbance of treated cells/Absorbance of control cells)×100. Cytotoxic concentra-
tion (IC50) of the samples was calculated by the dose–response curves. The antican-
cer data are mean standard deviation and were taken as the average of three sets of
readings of independent trials by taking doxorubicin as reference drug.
Synthesis
Synthesis of hydrazones (HL1–HL4) (1–4)
The
hydrazones:
HL2/4-methyl-benzoic
nicotinic
acid
acid
(3,4-dihydro-2H-naphthalen-1-ylidene)-
hydrazide HL1/benzoic acid (3,4-dihydro-2H-naphthalen-1-ylidene)-hydrazide
(3,4-dihydro-2H-naphthalen-1-ylidene)-hydrazide
HL3/isonicotinic acid (3,4-dihydro-2H-naphthalen-1-ylidene)-hydrazide HL4
were synthesized by the condensation of tetralone (0.438 g, 3.0 mmol) with nic-
otinic (0.411 g) HL1/benzoic (0.408 g) HL2/p-toluic (0.450 g) HL3/isonicotinic
(0.411 g) HL4 acid hydrazide (3.0 mmol) under refuxing in ethanol (20 mL) for
1 3

Transition metal (II) complexes of hydrazones derived from…
O
C
H
N
Ar
NH2
Reflux 2-4 h Ar
MeOH
N
+
tutomerism
N
H
C
N
Ar
C
O
N
OH
O
N
L1
=
5, 9, 13, 17
=
[Co(L1-4)2(H2O)2]
=
Ar
1.
6, 10, 14, 18 = [Ni(L1-4)2(H2O)2]
2. L2 = Ar =
H3C
Ar
N
C
O
N
M
N
3. L3 = Ar =
7, 11, 15, 19 = [Cu(L1-4)2(H2O)2]
8, 12, 16, 20 = [Zn(L1-4)2(H2O)2]
H2O
OH2
Ar
4. L4 = Ar =
N
O
C
N
M= Co(II), Ni(II), Cu(II) and Zn(II)
Scheme 1 Synthesis of hydrazones (1–4) and their transition metal complexes (5–20)
2–4 h in equimolar ratio under constant stirring. The reaction progress was regularly
monitored by TLC. The light colour solid compounds were separated by fltration,
washed with methanol, dried under vacuum over anhydrous calcium chloride, and
recrystallized from chloroform to obtain pure hydrazones (Scheme 1).
Nicotinic acid (3,4‑dihydro‑2H‑naphthalen‑1‑ylidene)‑hydrazide (HL1)
Yield: 87%; Colour: Light yellow; M.p.: 75–78 °C; Conductivity:
(ohm−1cm2 mol−1) in DMSO: 12; C16H15N3O: Anal. Cald: C, 72.43; H, 5.70;
N, 15.84, Found: C, 72.42; H, 5.73; N, 15.87; MS: m/z (M+) Cald: 265.31;
Found:266.7077 (M + 1)+; FT-IR (KBr, ν, cm−1): 1653 ν(C=O, carbonyl), 3258
ν(–NH),1615 ν(–C=N), 1013 ν(–N–N–H); 1H NMR [400 MHz, CDCl3, δ(ppm)]:
′
9.26 (s, t, 1H, N=C–OH), 7.85 (d, J = 4 Hz, 1H, C5 –H), 7.51 (d, J = 8 Hz, 1H,
C9′–H), 7.21 (d, J = 8 Hz, 2H, C6–H), 7.18 (d, J = 8 Hz, 2H, C7–H), 8.78 (s, 1H,
′
C2′–H), 7.36 (t, J = 8 Hz, 1H, C8–H), 7.25 (t, J = 4 Hz, 1H, C4 –H), 8.30 (s, 2H,
C3 –H), 2.85 (t, J = 6 Hz, 2H, C4–H), 2.71 (t, J = 6 Hz, 2H, C2–H), 2.04 (m,
J = 6 Hz, 2H, C3–H);13C NMR:[100 MHz, CDCl3, δ(ppm)]: 166.38 (HNC=O),
′
′
′
151.70 (C1′), 139.66 (C2 ′), 131.95 (C5 ), 129.94 (C9), 137.92 (C5), 122.92 (C4 ),
133.90 (C3 ), 133.60 (C1 ), 128.93 (C8), 126.61 (C6), 124.21 (C7), 120.12 (C10),
29.93 (C4), 24.58 (C2), 21.27 (C3).
1 3

J. Devi et al.
Benzoic acid (3,4‑dihydro‑2H‑naphthalen‑1‑ylidene)‑hydrazide (HL2)
Yield: 82%; Colour: white; M.p.: 72–75 °C; Conductivity: (ohm−1cm2mol−1)
in DMSO: 11, C17H16N2O: Anal. Cald: C, 77.25; H, 6.10; N, 10.60, Found.:
C, 77.22; H, 6.08; N, 10.58, MS: m/z (M+) Cald: 264.12; Found:265.1241
(M + 1)+; FT-IR (KBr, ν, cm−1): 1648 ν(C=O, carbonyl), 3242 ν(–NH),1612
1
1
ν(–C=N), 1013 ν(–N–N–H); H NMR [400 MHz, CDCl3, δ(ppm)]: 9.01 (s, H,
′
′
N=C–OH), 8.38 (s, 1H, C9–H), 7.89 (d, 2H, J = 12 Hz, C2 –H and C6 –H), 7.59
′
′
′
(t, J = 12 Hz, 1H, C4 –H), 7.52 (t, J = 12 Hz, 2H, C3 –H and C5 –H), 7.28 (d,
1H, C8–H. J = 8 Hz), 7.16 (d, J = 8 Hz,2H, C6–H and C7–H), 2.85 (t, J = 6 Hz,
2H, C4–H), 2.68 (t, J = 6 Hz, 2H, C2–H), 2.03 (m, J = 6 Hz, 2H, C3–H); 13C
NMR:[100 MHz, CDCl3, δ(ppm)]: 161.31 (HNC=O), 150.62 (C1), 139.63 (C5),
′
′
′
′
′
135.93 (C1′), 128.12 (C9), 131.27 (C2 ), 129.17 (C6 ), 133.61 (C4 ), 130.59 (C3 ),
127.88 (C5 ), 126.61 (C8), 125.55 (C6), 125.25 (C7), 121.18 (C10), 29.62 (C4),
25.25 (C2), 21.61 (C3).
4‑Methyl‑benzoic acid (3,4‑dihydro‑2H‑naphthalen‑1‑ylidene)‑hydrazide (HL3)
Yield: 79%; Colour: white; M.p.: 74–76 °C; Conductivity: (ohm−1cm2 mol−1)
in DMSO: 12; C18H18N2O: Anal. Cald: C, 77.67; H, 6.52; N, 10.06, Found.: C,
77.65; H, 6.50; N, 10.05, MS: m/z (M+) Cald: 278.15; Found:279.1575 (M + 1)+;
FT-IR (KBr, ν, cm−1): 1645 ν(C = O, carbonyl), 3246 ν(–NH),1610 ν(–C = N),
1015 ν(–N–N–H); 1H NMR [400 MHz, CDCl3, δ(ppm)]: 9.04 (s, 1H, N = C–OH),
′
′
8.37 (s, 1H, C9–H), 7.31 (d, J = 8.4 Hz, 2H, C2 –H and C6 –H), 7.28 (d, J = 8.4 Hz,
′
′
2H, C3 –H and C5 –H), 7.81 (t, J = 8 Hz, 1H, C8–H), 7.16 (d, J = 8 Hz, 2H, C6–H
and C7–H), 2.85 (t, J = 6.0 Hz, 2H, C4–H), 2.67 (t, J = 6 Hz, 2H, C2–H), 2.04 (m,
J = 6 Hz, 2H, C3–H), 2.45 (s, 3H, –CH3); 13C NMR: [100 MHz, CDCl3, δ(ppm)]:
′
163.63 (HNC=O), 153.64 (C1), 139.92 (C5), 132.54 (C1 ), 130.20, (C9), 127.90
′
′
′
′
′
(C2 ), 126.23 (C6 ), 133.58 (C4 ), 128.56 (C3 ), 127.58 (C5 ), 126.91 (C8), 124.57
(C6), 123.88 (C7), 122.24 (C10), 29.62 (C4), 23.29 (-CH3), 25.17 (C2), 21.58 (C3).
Isonicotinic acid (3,4‑dihydro‑2H‑naphthalen‑1‑ylidene)‑hydrazide (HL4)
Yield: 78%; Colour: white; M.p.: 77–79 °C; Conductivity: (ohm−1 cm2 mol−1)
in DMSO: 13.; C16H15N3O: Anal. Cald: C, 72.43; H, 5.71; N, 15.85, Found.: C,
72.42; H, 5.73; N, 15.87; MS: m/z (M+) Cald: 265.12; Found: 266.1277 (M + 1)+;
FT-IR (KBr, ν, cm−1): 1655 ν(C=O, carbonyl), 3248 ν(–NH),1611 ν(–C = N),
1
1
1018 ν(–N–N–H); H NMR [400 MHz, CDCl3, δ(ppm)]: 9.86 (s, H, N=C–OH),
′
′
8.24 (s, 1H, C9–H), 7.66 (d, J = 8 Hz, 2H, C2 –H and C5 –H), 8.67 (d, J = 8 Hz,
′
′
2H, C3 –H and C4 –H), 7.06 (t, J = 6.2 Hz, 1H, C8–H), 7.16 (d, J = 6.2 Hz, 2H,
C6–H and C7–H), 2.85 (t, J = 6 Hz, 2H, C4–H), 2.71 (t, J = 6, Hz, 2H, C2–H),
2.04 (m, J = 6 Hz, 2H, C3–H); 13C NMR:[100 MHz, CDCl3, δ(ppm)]: 162.37
′
′
(HNC=O), 147.15 (C1), 142.99 (C5), 133.22(C1 ), 126.85(C9), 128.55 (C2 ),
′
′
′
131.26 (C3 ), 130.22 (C4 ), 128.23 (C5 ), 129.24 (C8), 127.20 (C6), 123.90 (C7),
121.17 (C10), 29.61 (C4), 26.79(C2), 21.58 (C3).
1 3

Transition metal (II) complexes of hydrazones derived from…
Synthesis of transition metal complexes [M(L1–4)2(H2O)2] (5–20)
To the methanolic solution (20 mL) of metal acetates [M(CH3COO)2·xH2O]
of cobalt (0.248 g, 1.0 mmol)/nickel (0.248 g, 1.0 mmol)/copper (0.198 g,
1.0 mmol)/zinc (0.220 g, 1.0 mmol) was added a hot aqeuous methanolic solu-
tion (20 mL) of hydrazones HL1 (0.530 g, 2.0 mmol)/HL2( 0.528 g, 2.0 mmol)/
HL3 (0.556 g, 2.0 mmol)/HL4 (0.530 g, 2.0 mmol) with constant stirring in 1:2
molar ratio for 2–3 h. To maintain the pH of the solution, NaOH was added to it.
The separated compounds were fltered, recrystallized with methanol and washed
with hexane to separate out unreacted metal acetates to obtain pure product
(Scheme 1).
Co(L1)2 (H2O)2]
Yield: 73%; Colour: Reddish brown; M.p.: 180–85 °C; Conductivity:
(ohm−1 cm2 mol−1) in DMSO: 17; C32H32CoN6O4; Anal. Cald: C, 61.63; H,
5.18; N, 13.45, Co, 9.46;Found.: C, 61.64; H, 5.17; N, 13.48; MS: m/z (M+)
Cald: 623.5982; Found:624.5712 (M + 1)+; FT-IR (KBr, ν, cm−1): 1253 ν(C–O),
3427 ν(coordinated water),1598 ν(–C=N), 1018 ν(–N–N–H), 545 ν(Co–O), 435
ν(Co–N).
[Ni(L1)2(H2O)2]
Yield: 75%; Colour: Green; M.p.: 197–99 °C; Conductivity: (ohm−1 cm2 mol−1)
in DMSO: 12; C32H32NiN6O4; Anal. Cald: C, 61.69; H, 5.48; N, 13.43, Ni, 9.40;
Found.: C, 61.66; H, 5.17; N, 13.48; MS: m/z (M+) Cald: 623.3573; Found:
624.3307 (M+1)+; FT-IR (KBr, ν, cm−1): 1245 ν(C-O), 3425 ν(coordinated
water),1594 ν(–C=N), 1021 ν(–N–N–H), 547 ν(Ni–O), 440 ν(Ni–N).
[Cu(L1)2(H2O)2]
Yield: 78%; Colour: Brown; M.p.: 194–97 °C; Conductivity: (ohm−1 cm2 mol−1) in
DMSO: 18; C32H32CuN6O4; Anal. Cald: C, 61.19; H, 5.13; N, 13.35, Cu, 10.09;
Found.: C, 61.18; H, 5.13; N, 13.38, MS: m/z (M+) Cald: 628.2035; Found:629.1889
(M+1)+; FT-IR (KBr, ν, cm−1): 1248 ν(C-O), 3428 ν(coordinated water),1595
ν(–C=N), 1023 ν(–N–N–H), 543 ν(Cu–O), 442 ν(Cu–N).
[Zn(L1)2(H2O)2]
Yield: 76%; Colour: Reddish brown; M.p.: 192–95 °C; Conductivity:
(ohm−1 cm2 mol−1) in DMSO: 19; C32H32ZnN6O4; Anal. Cald: C, 61.02; H, 5.11;
N, 13.31, Zn:10.38; Found.: C, 61.00; H, 5.12; N, 13.34; MS: m/z (M+) Cald:
630.0549; Found: 631.0265 (M+1)+; FT-IR (KBr, ν, cm−1): 1233 ν(C–O), 3430
ν(coordinated water),1586 ν(–C=N), 1029 ν(–N–N–H), 548 ν(Zn–O), 439 ν(Zn–N),
′
1H NMR [400 MHz, CDCl3, δ(ppm)]: 7.85 (d, J=4 Hz, 1H, C5 –H), 7.51 (d,
1 3

J. Devi et al.
′
′
J=8 Hz, 1H, C9–H), 8.52 (d, J=8 Hz, 1H, C2 –H), 7.25 (t, J=4 Hz, 1H, C4 –H),
′
8.07 (s, 2H, C3 –H); 7.35 (t, J=8 Hz, 1H, C8–H), 7.21 (d, J=8 Hz, 2H, C6–H),
7.18 (d, J=8 Hz, 2H, C7–H), 2.86 (t, J=6.2 H, 2H, C4–H), 2.70 (t, J=5.2 Hz, 2H,
C2-H), 2.44 (s, 2H, H2O), 2.04 (m, J=6.8 Hz, 2H, C3-H); 13C NMR:[ 100 MHz,
′
′
CDCl3, δ(ppm)]: 169.02 (N=C–OH), 152.79 (C1), 139.60 (C2 ), 131.93 (C5 ), 129.84
′
′
′
(C9), 137.94 (C5), 122.92 (C4 ), 133.91 (C3 ), 133.62 (C1 ), 128.97 (C8), 126.62 (C6),
124.22 (C7), 120.16 (C10), 29.96 (C4), 24.23 (C2), 21.35(C3).
[Co(L2)2(H2O)2]
Yield: 77%; Colour: Reddish brown; M.p.: 195–97 °C; Conductivity:
(ohm−1 cm2 mol−1) in DMSO: 16; C34H34CoN4O4; Anal. Cald: C, 65.72; H, 5.51; N,
9.04; Co, 9.45; Found.: C, 65.70; H, 5.51; N, 5.01, MS: m/z (M+) Cald: 621.3527;
Found:622.6552 (M+1)+; FT-IR (KBr, ν, cm−1): 1249 ν(C–O), 3417 ν(coordinated
water),1591 ν(–C=N), 1016 ν(–N–N–H), 540 ν(Co–O), 426 ν(Co–N).
[Ni(L2)2(H2O)2]
Yield: 79%; Colour: Green; M.p.: 227–32 °C; Conductivity: (ohm−1 cm2 mol−1)
in DMSO: 14; C34H34NiN4O4; Anal. Cald: C, 65.71; H, 5.51; N, 9.04; Ni, 9.43;
Found.: C, 65.72; H, 5.52; N, 9.02; MS: m/z (M+) Cald: 621.5978; Found: 622.6252
(M+1)+; FT-IR (KBr, ν, cm−1): 1267, ν(C-O), 3415 ν(coordinated water),1588
ν(–C=N), 1017 ν(–N–N–H), 542 ν(Ni–O), 422 ν(Ni–N).
[Cu(L2)2(H2O)2]
Yield: 72%; Colour: Brown; M.p.: 216–19 °C; Conductivity: (ohm−1cm2 mol−1)
in DMSO: 17; C34H34CuN4O4; Anal. Cald: C, 65.23; H, 5.48; N, 8.92,Cu, 10.12;
Found.: C, 65.21; H, 5.47; N, 8.92; MS: m/z (M+) Cald: 627.2047; Found:628.2396
(M+1)+; FT-IR (KBr, ν, cm−1): 1248 ν(C–O), 3422 ν(coordinated water),1589
ν(–C=N), 1018 ν(–N–N–H), 539 ν(Cu–O), 425 ν(Cu–N).
[Zn(L2)2(H2O)2]
Yield: 75%; Colour: Reddish brown; M.p.: 218–22 °C; Conductivity:
(ohm−1 cm2 mol−1) in DMSO: 20; C34H34ZnN6O4; Anal. Cald: C, 65.04; H,
5.45; N, 8.92, Zn; 10.38; Found.: C,65.02; H,5.46; N, 8.91; MS: m/z (M+) Cald:
628.0767; Found:629.0546 (M+1)+; FT–IR (KBr, ν, cm−1): 1253 ν(C–O), 3425,
ν(coordinated water),1585 ν(–C=N), 1019 ν(–N–N–H), 537 ν(Zn–O), 427 ν(Zn–N);
1H NMR [400 MHz, CDCl3, δ(ppm)]: 8.32 (d, 1H, C9–H), 7.88 (d, J=12, Hz 2H,
′
′
′
′
C2′–H and C6 –H), 7.61 (t, J=12 Hz, 1H, C4 –H), 7.52 (t, J=12 Hz, 2H, C3 –H and
C5 –H), 7.28 (d, J=8 Hz, 1H, C8–H), 7.16 (d, J=8 Hz, 2H, C6–H and C7–H), 2.84
(t, J=6 Hz, 2H, C4–H), 2.68 (t, J=6 Hz, 2H, C2–H), 2.03 (m, J=6 Hz, 2H, C3–H),
1 3

Transition metal (II) complexes of hydrazones derived from…
2.40 (s, 2H, H2O); 13C NMR:[100 MHz, CDCl3, δ(ppm)]: 164.02 (N=C–OH),
′
′
′
153.63(C1), 142.26 (C5), 135.95(C1 ), 128.54 (C9), 131.94 (C2 ), 129.91 (C6 ), 133.23
′
′
′
(C4 ), 130.60 (C3 ), 127.58 (C4 ), 126.61 (C8), 125.55 (C6), 125.25 (C7), 121.19 (C10),
29.63 (C4), 25.23 (C2), 20.28 (C3).
[Co(L3)2(H2O)2]
Yield: 73%; Colour: Reddish brown; M.p.: 178–181 °C; Conductivity:
(ohm−1cm2 mol−1) in DMSO: 13; C36H38CoN4O4; Anal. Cald: C: 66.55; H: 5.92; N,
8.65, Co, 9.04; Found.: C, 66.56; H, 5.90; N, 8.62; MS: m/z (M+) Cald: 649.6586;
Found:650.2863 (M+1)+; FT-IR (KBr, ν, cm−1): 1262, ν(C–O), 3431 ν(coordinated
water),1599 ν(–C=N), 1021 ν(–N–N–H), 547 ν(Co–O), 434, ν(Co–N).
[Ni(L3)2(H2O)2]
Yield: 75%; Colour: Green; M.p.: 214–218 °C; Conductivity: (ohm−1 cm2 mol−1)
in DMSO: 16; C36H38NiN4O4; Anal. Cald: C, 66.57; H, 5.91; N, 8.60,,Ni, 9.01;
Found.: C, 66.58; H, 5.90; N, 8.63; MS: m/z (M+) Cald: 649.4219.; Found:650.4258
(M+1)+; FT-IR (KBr, ν, cm−1): 1225 ν(C–O), 3434, ν(coordinated water),1590
ν(–C=N), 1023 ν(–N–N–H), 540 ν(Ni–O), 435 ν(Ni–N).
[Cu(L3)2(H2O)2]
Yield: 78%; Colour: Brown; M.p.: 192–96 °C; Conductivity: (ohm−1cm2 mol−1)
in DMSO: 17; C36H38CoN6O4; Anal. Cald: C, 66.07; H, 5.84; N, 8.56;Cu, 9.69;
Found.: C, 66.09; H, 5.85; N, 8.51; MS: m/z (M+) Cald: 654.3189; Found:655.4115
(M+1)+; FT-IR (KBr, ν, cm−1): 1234 ν(C–O), 3437 ν(coordinated water),1591
ν(–C=N), 1025 ν(–N–N–H), 554 ν(Cu–O), 437 ν(Cu–N).
[Zn(L3)2(H2O)2]
Yield: 76%; Colour: white; M.p.: 193–93 °C; Conductivity: (ohm−1 cm2 mol−1)
in DMSO: 18; C36H38CoN6O4; Anal. Cald: C, 65.92; H, 5.83; N, 8.51, Zn:9.71;
Found: C, 65.90; H, 5.84; N, 8.54; MS: m/z (M+) Cald: 656.1274; Found:657.1084
(M+1)+; FT-IR (KBr, ν, cm−1): 1255 ν(C–O), 3435 ν(coordinated water),1592
1
ν(–C=N), 1024 ν(–N–N–H), 550 ν(Zn–O), 431 ν(Zn–N); H NMR [400 MHz,
′
CDCl3, δ(ppm)]:. 8.21(d, J=8 Hz, 1H, C9–H), 7.30 (t, J=8.4 Hz, 1H, C4 –H and
′
′
′
C2 –H), 7.28 (d, J=8.4 Hz, 2H, C3 –H and C5 –H), 7.62 (t, J=8 Hz, 1H, C8–H), 7.16
(d, J=8 Hz, 2H, C6–H and C7–H), 2.85 (t, J=6 Hz, 2H, C4–H), 2.67 (t, J=6 Hz,
2H, C2–H), 2.03 (m, J=6 Hz, 2H, C3–H), 2.46 (s, 3H, –CH3), 2.38 (s, 2H, H2O);
13C NMR:[100 MHz, CDCl3, δ(ppm)]: 165.37 (N=C–OH), 154.62 (C1), 143.61
′
′
′
′
(C5′), 132.54 (C1 ), 130.20, (C9), 127.90 (C2 ), 126.23 (C6 ), 133.58 (C4 ), 128.56
′
′
(C3 ), 129.74 (C4 ), 127.98 (C5 ), 126.91 (C8), 124.44 (C6), 123.86 (C7), 122.25 (C10),
29.62 (C4), 23.19 (–CH3), 25.27 (C2), 21.58 (C3).
1 3

J. Devi et al.
[Co(L4)2(H2O)2]
Yield: 75%; Colour: Reddish brown; M.p.: 213–16 °C; Conductivity:
(ohm−1cm2 mol−1) in DMSO: 15; C32H32CoN6O4; Anal. Cald: C, 61.61; H, 5.15; N,
13.49, Co, 9.43; Found.: C, 61.64; H, 5.17; N, 13.48; MS: m/z (M+) Cald: 625.1068;
Found:626.1076 (M+1)+; FT-IR (KBr, ν, cm−1): 1253 ν(C–O), 3437 ν(coordinated
water),1588 ν(–C=N), 1018 ν(–N–N–H), 545 ν(Co–O), 442 ν(Co–N).
[Ni(L4)2(H2O)2]
Yield: 74%; Colour: Green; M.p.: 242–46 °C; Conductivity: (ohm−1cm2mol−1) in
DMSO: 17; C32H32NiN6O4; Anal. Cald: C, 61.67; H, 5.20; N, 13.49, Ni, 9.44; Found.:
C, 61.66; H, 5.17; N, 13.48; MS: m/z (M+) Cald: 625.1768; Found:626.3393 (M+1)+;
FT-IR (KBr, ν, cm−1): 1232 ν(C–O), 3440 ν(coordinated water),1584 ν(–C=N), 1021
ν(–N–N–H), 546 ν(Ni–O), 436 ν(Ni–N).
[Cu(L4)2(H2O)2]
Yield: 77%; Colour: Brown; M.p.: 223–28 °C; Conductivity: (ohm−1cm2 mol−1) in
DMSO: 18; C32H32CoN6O4; Anal. Cald: C, 61.20; H, 5.15; N, 13.37,Cu, 10.14; Found.:
C, 61.18; H, 5.13; N, 13.38; MS: m/z (M+) Cald: 628.1557; Found:629.4533 (M+1)+;
FT-IR (KBr, ν, cm−1): 1228 ν(C–O), 3448 ν(coordinated water),1593 ν(–C=N), 1023
ν(–N–N–H), 543 ν(Cu–O), 440 ν(Cu–N).
[Zn(L4)2(H2O)2]
Yield: 79%; Colour: white; M.p.: 230–33 °C; Conductivity: (ohm−1 cm2 mol−1)
in DMSO: 19; C32H34CoN6O4; Anal. Cald: C, 61.03; H, 5.14; N, 13.33, Zn: 10.40;
Found.: C, 61.00; H, 5.12; N, 13.34; MS: m/z (M+) Cald: 630.0523; Found:631.1550
(M+1)+; FT-IR (KBr, ν, cm−1): 1253 ν(C–O), 3446 ν(coordinated water),1596
1
ν(–C=N), 1029 ν(–N–N–H), 547 ν(Zn–O), 445 ν(Zn–N); H NMR [400 MHz,
′
′
CDCl3, δ(ppm)]: 8.03 (s, 1H, C9–H), 7.66 (d, J=8 Hz, 2H, C2 –H and C5 –H), 8.66 (d,
′
′
J=8 Hz, 2H, C3 –H and C4 –H), 7.06 (t, J=6.2 Hz, 1H, C8–H), 7.16 (d, J=6.2 Hz, 2H,
C6–H and C7–H), 2.85 (t, J=6 Hz, 2H, C4–H), 2.68 (t, J=6 Hz, 2H, C2–H), 2.04 (m,
J=6 Hz, 2H, C3–H), 2.40 (s, 2H, H2O); 13C NMR:[100 MHz, CDCl3, δ(ppm)]: 166.34
′
′
(N=C–OH), 149.63 (C1), 142.95(C5), 133.22(C1 ), 126.85(C9), 128.55 (C2 ), 131.26
′
′
′
(C3 ), 130.22 (C4 ), 128.23 (C5 ), 129.24 (C8), 127.20 (C6), 123.90 (C7), 121.17 (C10),
29.61 (C4), 26.31(C2), 21.28 (C3).
Results and discussion
The hydrazones were synthesized by the condensation of respective hydrazide deriva-
tive with tetralone in equimolar ratio under refuxing in ethanol, and their transition
metal complexes were obtained by reacting with Co(II), Ni(II), Cu(II) and Zn(II)
1 3

Transition metal (II) complexes of hydrazones derived from…
1 3

J. Devi et al.
acetates at constant stirring in molar ratio 2:1. All the compounds were soluble in
MeOH, CDCl3 and DMSO but were insoluble in water. The hydrazones and transi-
tion metal complexes (1–20) were characterized by various spectroscopic techniques
(NMR, Mass, FT-IR, ESR, electronic, fuorescence), which suggests octahedral geom-
etry of complexes formulated as [M(L1−4)2(H2O)2] where hydrazones get bonded to
metal centre through the nitrogen of (C=N) and oxygen of carbonyl group in enolic
form. The compounds were stable in air, diferently coloured solid, obtained in good
yield and non-electrolytic in nature as their molar conductance values are in range
11–20 Ω−1cm2mol−1 [31]. The analytical data are depicted in experimental part and in
Table 1 given in supplementary data.
(
)
(
) (
)
M CH3COO ⋅ xH2O + HL1−4 → [M L1−4 H2O ] + 2CH3COOH
2
2
2
where M=Co(II), Ni(II), Cu(II) and Zn(II).
IR spectra of compounds
The FT-IR spectral analysis of compounds was obtained in 400–4000 cm−1 region
by using KBr pellets. The bands of hydrazones and their respective complexes tabu-
lated in Table 2 given in supplementary data and experimental part were compared
with each other to confrm the coordination sites on the basis of the shifts in the
Table 2 Characteristic IR frequencies ν(cm−1) of hydrazones and their respective transition metal(II)
complexes
C. no. Compounds
ν(–HOH)
ν(C=O)
ν(–C=N)
ν(N–H) ν(N–N–H)
ν(M–O) ν(M–N)
water
amide
1
2
3
4
5
6
7
8
HL1
HL2
HL3
HL4
–
–
–
–
1653
1648
1645
1655
–
–
–
–
–
–
–
–
–
–
–
–
–
1615
1612
1610
1613
1598
1594
1595
1586
1591
1588
1589
1585
1599
1590
1591
1592
1588
1584
1593
1596
3258
3242
3246
3248
–
–
–
–
–
–
–
–
–
–
–
–
–
1013
1011
1015
1018
1017
1021
1023
1029
1016
1017
1018
1019
1021
1023
1025
1024
1024
1021
1026
1025
–
–
–
–
–
–
–
–
[Co(L1)2(H2O)2] 3427
[Ni(L1)2(H2O)2]
3425
[Cu(L1)2(H2O)2] 3428
[Zn(L1)2(H2O)2]
3430
[Co(L2)2·(H2O)2] 3417
[Ni(L2)2(H2O)2]
3415
[Cu(L2)2(H2O)2] 3422
[Zn(L2)2(H2O)2]
3425
[Co(L3)2(H2O)2] 3431
[Ni(L3)2(H2O)2]
3434
[Cu(L3)2(H2O)2] 3437
545
547
543
548
540
542
539
537
547
540
554
550
545
546
543
547
435
440
442
439
426
422
425
427
434
435
437
431
442
436
440
445
9
10
11
12
13
14
15
16
17
18
19
20
[Zn(L3)2(H2O)2]
3435
[Co(L4)2(H2O)2] 3437
[Ni(L4)2·(H2O)2] 3440
[Cu(L4)2(H2O)2] 3448
–
–
–
–
–
–
[Zn(L4)2(H2O)2]
3446
1 3

Transition metal (II) complexes of hydrazones derived from…
absorbance frequency of diferent types of groups and the absence of certain absorp-
tion band due to changes in the electronic environment of the hydrazones after com-
plexation. A strong band in ligands at 1615–1610 cm−1 due to azomethine group
shifts to the lower-frequency region 1599–1584 cm−1 on complexation with the
metal atom; this downfeld shifting shows the bonding via lone pair of the nitrogen
of (C=N) group to metal centre [32, 33]. The bands in the region 1655–1645 cm−1
due to υ(C=O) group and at 3258–3242 cm−1 due to υ(N–H) group in Schif base
hydrazones got disappeared, indicating their enolization and chelation with metal
centre. The shifting of υ(N–N-H) band at 1018–1011 cm−1 in free hydrazone to
higher value 1029–1016 cm−1 in the transition metal complexes confrmed the
coordination via lone pair electron of the nitrogen to the metal centre. The appear-
ance of new sharp and distinct bands in the far-infrared spectra of all complexes
at 554–537 cm−1 and 445–422 cm−1, due to M–O and M–N bonds, suggests the
coordination of metal with oxygen and nitrogen atoms [34]. A broad peak near
3448–3415 cm−1 is due to stretching vibration of water molecules coordinated to
the central metal ion [35]. From the above IR data, it is suggested that hydrazones
are bidentate (NO) in nature, and they bind with the metal ion through a nitrogen of
(C=N) group and oxygen of carbonyl group.
NMR spectra of compounds
1H NMR spectra
3
3
1
3'
4
3'
4
3'
3
3
4
H3C
4'
5'
2'
1'
2'
1'
N
5'
2'
1'
4
2'
1'
3'
4'
4'
5'
2
5
2
2
5
N
4'
2
10
5
5
8
10
6
10
6
7
6
6
7
N
1
N
N
N
C
N
7
1
C
C
N
C
N
N
7
1
10
9
6'
6'
5'
9
8
9
OH
9
8
OH
HL3
OH
HL1
8
OH
HL4
HL2
NMR spectra of hydrazones and their zinc(II) complexes were obtained in CDCl3,
and their δ values are presented in "experimental" section and Table 3 given in sup-
plementary data. The binding sites (NO) of the hydrazones were confrmed by cor-
relating spectra of ligands with respective zinc complexes. The hydrazones (1–4)
exhibit a singlet at δ 9.86–8.88 ppm due to (–NH) proton which gets disappeared
upon complexation, and the proton loss on the nitrogen indicated deprotonation
of –NH group due to the presence of keto-enol tautomerism on coordination with
the transition metal ion. The metal binds through nitrogen of (C=N) and oxygen
of deprotonated carbonyl oxygen in enolized form showing bidentate nature of
hydrazones, which was also supported with the IR spectra results [36]. The aro-
matic protons were recognized in the range δ 8.78–7.06 ppm, which exhibits only
minor change in the complexes as a result of binding with the metal ion [37]. A
sharp singlet of the three hydrogen atoms of the methyl group at 2.45 ppm in HL3
ligand remains almost unaltered on complexation. The signals due to CH2 group of
1 3

J. Devi et al.
1 3

Transition metal (II) complexes of hydrazones derived from…
1 3

J. Devi et al.
aliphatic ring protons of C2, C3, and C4 carbon are in the range δ 2.85–2.04 ppm,
which stands almost unaltered on complexation.
13C NMR spectra
The 13C NMR spectra of hydrazones showed that characteristic signals at δ
166.38–161.31 ppm due to (HNC=O) carbon, which display upward shifting in
the complexes, indicated the coordination of oxygen atom through lone pair elec-
trons of carbonyl group to the metal ion and the upfeld shifting of C1 carbon of
(C=N) group from δ 153.64–147.15 to δ 154.62–149.63 ppm reveals the coordina-
tion of lone pair of nitrogen to the metal ion centre. The peaks of aromatic carbons
in hydrazones were found near δ 142.99–120.12 ppm, which exhibited only small
alteration upon coordination with transition metal ion. The signal due to carbon of
(–CH3) in HL3 ligand was observed near δ 23.29 ppm and remains unaltered upon
complexation. The signals at δ 29.93–21.27 ppm due to the aliphatic ring C2, C3,
and C4 carbons remain almost unchanged. All peak values (1H NMR and 13C NMR)
and their graph related to synthesized ligands and their Zn(II) complexes are given
in supplementary data.
Mass spectra of compounds
The mass spectra of the hydrazones and their metal complexes were obtained and
are specifed in analytical Table 1, and their molecular ion peaks are in agreement
with the expected molar weight values. The mass spectrum provides valuable infor-
mation about the molar weight and structure of compounds. The mass fragmenta-
tion of the ligand HL4 (4) is depicted in Fig. 1 and Scheme 2. The mass spectra
show that a molecular ion peak of high intensity at m/z=266.1277 is assigned for
[M+1]+, which shows two fragmentation paths where one path shows a peak value
at m/z=189.1113 due to loss of pyridine moiety followed by mass spectrum peak at
m/z=161.0964 due to loss of CO group and at m/z=145.0761 by loss of NH group.
Another fragmentation path depicts peaks at m/z=139.0648 due to loss of pyridine
ring and aromatic ring followed by m/z=111.0568 due to loss of CO group.
In the mass spectra of [Co(L4)2(H2O)2] (17) the molecular ion peak at
m/z=626.1076 due to [M+1]+ is observed, which agrees with its molecular weight
which are described in Fig. 2. The molecular ion peak obtained at m/z=587.1855 is
due to loss of both water molecules coordinated to the transition metal. The peak at
m/z 266.1624 is due to loss of one ligand moiety and cobalt metal itself (Scheme 3)
and after this complex follows the similar fragmentation pattern as hydrazone HL4.
Based upon the above mass spectral results, all the transition metal complexes
follow the similar fragmentation model. The molecular ion peaks obtained from
mass spectra were found to be in good agreement with the expected values of the
elemental analysis. The mass spectra of all the hydrazones and their metal com-
plexes indicate their bidentate nature, and the complexation of hydrazones with cen-
tral metal atom is in 2:1 ratio with molecular formula [M(L1−4)2(H2O)2].
1 3

Transition metal (II) complexes of hydrazones derived from…
Fig. 1 Mass spectra of hydrazone HL4 (4)
1 3

J. Devi et al.
N
N
NH
-
C
O
N
N
,
+
-
,
tutomerism
H
NH
+
N
C
O
N
N
C
N
H
NH
,
C
11H12N2O
OH
C
O
N
m/z =189.1113 (M+1)
C7H10N2O
m/z = 139.0646 (M+1)
C16H15N3O
m/z = 266.1277(M+1)
-CO
-CO
+
+
NH
H
N
H
N
H
N
C10H12N2
m/z =161.0954 (M+1)
C6H10N2
m/z =111.0586 (M+1)
-NH
+
H
N
C
10H11
N
m/z = 145.0764 (M+1)
Scheme 2 The possible molecular ion fragmentation pattern from the mass spectrum of hydrazone HL4
(4)
ESR spectra of compounds
The solid-state ESR analysis of the copper (II) complexes was obtained at RT
(300 K) and LNT (77 K) in DMSO to gather information about stereochemis-
try, bonding nature of ligands and metal ion. From the ESR data, the spin Ham-
iltonian parameters were determined and are summarized in Table 4. The ESR
spectrum of [Cu(L3)2(H2O)2] (15) complex describes a strong band in the upfeld
region, which may be due to tumbling. The trend followed for g tensor values was:
g|| (2.43)>g⊥(2.14)>gc(2.0023), suggesting residence of single unpaired electron in
2
the dx −y2 orbital of Cu(II) ions in the ground state [38], which is predictable feature
of octahedral geometry around metal ions and gav was obtained by the formula: 1/3
1 3

Transition metal (II) complexes of hydrazones derived from…
Fig. 2 Mass spectra of [Co(L4)2(H2O)2] (17)
(g||+2 g⊥) which were found to be lower than free copper(II) ion λ (832 cm−1). Spin
orbital coupling constant λ is calculated from relation gav =2(1−2 λ/10Dq), and it
supports the evidence for metal→ligand bond to be covalent in nature, and this was
also confrmed from gav values (2.23) of the complex and the spectral parameters of
Cu(II) complexes are given in Table 4.
The measured anisotropic geometric parameter G and g tensor values indicate the
exchange interactions among the copper(II) ion and may be calculated by utilizing
the subsequent equation G=(g||−2.0023)/(g⊥ −2.0023) [39]. If G<4, then diferent
types of exchange interactions among the Cu(II) centre are present, and if G>4,
then these exchange interactions were minor. In all these copper complexes, the G
1 3

J. Devi et al.
+
+
N
O
N
N
N
N
N
O
H
H
OH2
-2H2O
Co
Co
H2O
O
N
O
N
N
N
N
N
C32H32CoN6O4
m/z = 626.1076 (M+1) peak
C32H30CoN6O2
m/z = 587.1825 (M+1) peak
- one ligand moiety+ Co
HO
N
O
N
N
N
N
NH
C15H15CoN3O
m/z = 266.1624 (M+1) peak
Scheme 3 The possible molecular ion fragmentation pattern for the complex [Co(L4)2(H2O)2] (17)
values were found to be in range 2.96–3.69, indicative of considerable exchange
interactions among the Cu(II) centres in the solid steady state [40].
Electronic spectra of compounds
The electronic spectra of the compounds were carried out in solvent DMSO, and
their band assignments are given in Table 5. The electronic spectra of free hydra-
zones showed a band at 28,278–29,815 cm−1 and 31,673–33,745 cm−1 attributed
to n→π* and π→π* transitions. Shifting in the value of n→π* transition shows
Table 4 ESR spectral
Cu(II) complexes
g||
g⊥
gav
G
parameters of Cu(II) complexes
[Cu(L1)2(H2O)2]
[Cu(L2)2(H2O)2]
[Cu(L3)2(H2O)2]
[Cu(L4)2(H2O)2]
2.40
2.43
2.41
2.44
2.11
2.14
2.12
2.15
2.20
2.23
2.21
2.24
3.69
3.10
3.46
2.96
1 3

Transition metal (II) complexes of hydrazones derived from…
Table 5 Electronic spectral data and magnetic moment (BM) of hydrazones and their transition metal(II)
complexes
Compounds Ligand/Complex Band assignment Transitions
μef(BM) Geometry
1
2
3
4
5
[HL1]
[HL2]
[HL3]
[HL4]
31,698
29,247
31,673
28,278
33,745
29,815
32,689
29,231
π→π*
n→π*
π→π*
n→π*
π→π*
n→π*
π→π
n→π*
–
–
–
–
–
–
–
–
[Co(L1)2(H2O)2] 23,531
4T1g(F)→4T1g(P) (v1)
4T1g(F)→4A2g(F) (v2)
4T1g(F)→4T2g(F) (v3)
3A2g(F)→3T2g(F) (v1)
3A1g(F)→3T1g(F) (v2)
3A2g(F)→3T1g(P) (v3)
2B1g →2A1g (v1)
2B1g →2E2g (v2)
LMCT
4.70
Octahedral
17,745
9577
24,345
18,332
10,467
6
7
[Ni(L1)2(H2O)2]
3.00
1.91
Octahedral
Octahedral
[Cu(L1)2(H2O)2] 24,156
15,823
8
9
[Zn(L1)2(H2O)2]
24,456
–
4.60
Octahedral
Octahedral
[Co(L2)2(H2O)2] 23,423
4T1g(F)→4T1g(P) (v1)
4T1g(F)→4A2g(F) (v2)
4T1g(F)→4T2g(F) (v3)
3A2g(F)→3T2g(F) (v1)
3A1g(F)→3T1g(F) (v2)
3A2g(F)→3T1g(P) (v3)
2B1g →2A1g (v1)
2B1g →2E2g (v2)
LMCT
17,734
9455
24,291
18,226
10,453
10
11
[Ni(L2)2(H2O)2]
2.80
1.78
Octahedral
Octahedral
[Cu(L2)2(H2O)2] 24,042
15,618
12
13
[Zn(L2)2(H2O)2]
24,440
–
5.00
Octahedral
Octahedral
[Co(L3)2(H2O)2] 23,649
4T1g(F)→4T1g(P) (v1)
4T1g(F)→4A2g(F) (v2)
4T1g(F)→4T2g(F) (v3)
3A2g(F)→3T2g(F) (v1)
3A1g(F)→3T1g(F) (v2)
3A2g(F)→3T1g(P) (v3)
2B1g →2A1g (v1)
2B1g →2E2g (v2)
LMCT
17,861
9587
24,675
18,645
10,582
14
15
[Ni(L3)2(H2O)2]
3.30
1.95
Octahedral
Octahedral
[Cu(L3)2(H2O)2] 24,272
15,847
16
17
[Zn(L3)2(H2O)2]
24,468
–
4.90
Octahedral
Octahedral
[Co(L4)2(H2O)2] 23,527
4T1g(F)→4T1g(P) (v1)
4T1g(F)→4A2g(F) (v2)
4T1g(F)→4T2g(F) (v3)
17,844
9575
24,437
18,429
10,664
18
[Ni(L4)2(H2O)2]
3A2g(F)→3T2g(F) (v1) 3.20
3A1g(F)→3T1g(F) (v2)
Octahedral
3A2g(F)→3T1g(P)(v3)
1 3

J. Devi et al.
Table 5 (continued)
Compounds Ligand/Complex Band assignment Transitions
μef(BM) Geometry
19
[Cu(L4)2(H2O)2] 24,214
2B1g →2A1g (v1)
1.85
Octahedral
15,720
24,450
2B1g →2E2g (v2)
LMCT
20
[Zn(L4)2(H2O)2]
–
Octahedral
the complexation of metal ion with hydrazones and the transitions due to aromatic
carbon (π→π*) remains unaltered. The spectra of the [Co(L1−4)2(H2O)2] com-
plexes exhibit three d–d absorption transitions in the range 23,423–23,649 cm−1,
4
17,734–17,861 cm−1, and 9455–9587 cm−1 due to T1g(F)→4T1g(P),
4
4T1g(F)→4A2g(F) and T1g(F)→4T2g(F) transitions which suggests octahedral
geometry of cobalt(II) complexes [41].
The ESR graph is depicted in Fig. 3 at room temperature and liquid nitrogen
temperature.
The spectra of [Ni(L1−4)2(H2O)2] complexes showed three d-d absorp-
tion transitions in the range of 24,291–24,675 cm−1, 18,226–18,645 cm−1 and
3
3
10,453–10,582 cm−1, ascribed to the A2g(F)→3T1g(P), A1g(F)→3T1g(F)
and A2g(F)→3T2g(F) transitions, respectively, which results in the octahe-
3
dral geometry of nickel(II) ions [42]. The spectra of [Cu(L1−4)2(H2O)2] com-
plexes showed two absorption transitions at 24,042–24,272, 15,618–15,847 cm−1
assigned to B1g →2A1g(ν1) and B1g →2E2g (v2) transitions [43], and spec-
tral data of [Zn(L1−4)2(H2O)2] complexes show only one band in the range of
24,440–24,468 cm−1 which is attributed to LMCT transitions [44], and these com-
plexes were diamagnetic in nature with complete flled d10 confguration. Based
upon the above spectral analysis and analytical data, it was confrmed that all the
complexes have octahedral arrangement around central metal ion [45].
2
2
Magnetic measurement of compounds
The magnetic moment data of Co(II), Ni(II) and Cu(II) complexes were found to be
in range of 4.60–5.00 BM, 2.80–3.30 BM and 1.78–1.95 BM, respectively, which
shows the paramagnetic nature of all the three types of metal complexes. The meas-
ured μef values are in the range known for six coordinated octahedral geometry [46]
and the Zn(II) complexes shows a diamagnetic behaviour.
Fluorescence spectra of compounds
The steady-state fuorescence emission spectral analysis of the hydrazones and
their corresponding metal complexes were carried out in CHCl3 with an excita-
tion wavelength of 300 nm at 298 K. On complexation, hydrazones indicate some
remarkable changes in fuorescent properties like quenching of spectra, variation in
intensity, diference of wavelength in emission bands or emergence of new bands,
etc. Hydrazone HL2 (2) observes broad fuorescent emission band at 407 nm, gets
1 3

Transition metal (II) complexes of hydrazones derived from…
Fig. 3 ESR spectrum of [Cu(L3)2(H2O)2](15) at a room temperature and b liquid nitrogen temperature
1 3

J. Devi et al.
Fig. 4 Fluorescence spectra of hydrazone HL2 (2) and its transition metal complexes
shifted on coordination with central metal atom and is shown in Fig. 4. The Co(II),
Ni(II), Cu(II) and Zn(II) metal complexes that exhibited bands at 410, 408, 412 and
415 nm, respectively, confrm complexation. The intensifcation in fuorescent spec-
tra (hyperchromic shift) of metal complexes can be ascribed for M→L charge trans-
fer (LMCT) transitions, which showed their extra stability [47]. The order followed
for fuorescent properties of hydrazone HL2 and its metal complexes follows the
trend: Zinc(II)>Copper(II)>Cobalt(II)>Nickel(II)>HL2. The enhancement in fu-
orescent intensity of zinc complexes is due to their fulflled d10 confguration mak-
ing them difcult to oxidize and reduce. Based on the above data, it was revealed
that the hydrazones and their transition metal complexes are good fuorescent com-
ponents and may have good photochemical applications [48].
Conductance measurements of compounds
The molar conductance of Co(II), Ni(II), Cu(II) and Zn(II) complexes of 1×10−3
M
solution is found to be between 11–20 Ω−1 cm2 mol−1, which confrmed the non-elec-
trolytic nature of compounds [31].
Thermal studies
Thermal behaviour of compounds was analysed by TG/DTA/DTG analysis, and
thermal data of the compounds were recorded between temperature 25–550 °C
1 3

Transition metal (II) complexes of hydrazones derived from…
Fig. 5 Thermo gravimetric curve of [Ni (L1)2(H2O)2] (6)
under nitrogen atmosphere with heating rate of 20 °C min−1. In the current study,
thermal analysis was carried out to get valuable information about the thermal sta-
bility of synthesized complexes and to decide whether water molecules are outside
or inside the coordination sphere of metal [49–51]. The various thermal decay steps
followed by the complexes with their respective mass loss are presented in Fig. 5.
The TG curve of all the complexes follows the same design of three-step decompo-
sition with mass loss till the metal oxide is formed. Thermal analysis describes that
the complexes are stable below 180 °C, which indicates that there are no hydrated
water molecules of crystallization as no mass loss was noticed up to this tempera-
ture, and the frst decay step is in the range 180–250 °C revealing the existence of
bonded water molecules to central metal atom. The second decay step takes place
between the temperature 250 °C and 350 °C with a mass loss attributed to a strong
broad exothermic peak on the TG graph agree with thermal decay process of the
one hydrazone moiety. The removal of second hydrazone moiety occurs in the range
350–550 °C. The thermal decay of all the transition metal complexes ended with
emergence of metal oxide residue[52, 53].
The three-step thermal decay process can be represented as:
[M (L1−4)2(H2O)2]→[M (L1−4)2(H2O)2] (no mass loss below 180 °C).
[M (L1−4)2(H2O)2]→[M (L1−4)2]+2H2O (Decomposition of two water mole-
cules, near 180–250 °C, endothermic peak).
[M (L1−4)2]→Intermediate [M (L1−4)] (Decomposition of one hydrazone moiety,
near 250–350 °C, exothermic peak).
Intermediate→Loss of other hydrazone moiety+metal oxide (Loss of another
hydrazone near 350–550 °C, exothermic peak).
Above 550 °C—metal oxide as a residue.
where M=Co(II), Ni(II), Cu(II) and Zn(II).
1 3

J. Devi et al.
The TG curve of the Ni(II) complex (6) (Fig. 5) revealed three thermal decay
stages in the range 193 to 560 °C and complex is highly stable up to 203 °C, which
indicates the absence of hydrated water molecules. The frst stage indicates a mass
loss of 5.7% (calculated 5.4%) in range 203–233 °C (DTG), which is due to the
removal of two H2O molecules with an endothermic peak at 226 °C. The second
stage takes place in the temperature range 233–350 °C with a mass loss of 42.18%
(calculated 42.02%) with the appearance of a strong broad exothermic peak exactly
at 325 °C. DTA and DTG curve reveal the thermal decay of one hydrazone moi-
ety. In the third stage, dropping of second hydrazone moiety occurs in this tempera-
ture interval 350–550 °C with a fall in mass of 42.18% (calculated 42.02%) with the
appearance of a strong broad exothermic peak exactly at 426 °C leaving nickel oxide
as residue at the end of thermal decay process [54]. The residual mass (12.3%) is
presumed to be nickel(II) oxide.
Biological studies
Antimicrobial activity
Hydrazones (HL1−4), their chelated transition metal complexes and standard drugs
were evaluated for antimicrobial activity (in vitro) using a serial dilution method
against two gram-positive bacteria (S. aureus, B. subtilis), 2 g negative bacteria (E.
coli, P. aeruginosa) and two fungal species (C. albicans, A. niger). Ciprofoxacin
and fuconazole drugs were used as a positive control for antibacterial and antifungal
activities, whereas DMSO is used as negative control having negligible inhibitory
efect. Minimum inhibitory concentration (MIC) in μmol/mL is depicted in Table 6
and its charted form is in Fig. 6.
Conclusion drawn from the biological data is summarized below:
1. The order of antimicrobial activity for hydrazones (HL1−4) was found as
HL3 >HL1 >HL4 >HL2, which is explained on the basis of nature of compounds
[55]. The antimicrobial evaluation of hydrazones is ascribed to the existence of
the azomethine (–C=N–) group, which have fne chelating abilities, which results
in construction of hydrogen bond with the active centres of the cell constituents,
which results in hindrance of normal life process [56]. The presence of electron
releasing methyl groups in hydrazone HL3 makes it highly active.
2. In antibacterial activity, complexes 15, 16, 19 having MIC values in the range
0.0091–0.0317 μmol/mL were found to be more or equally active than standard
ciprofoxacin (MIC=0.0188 μmol/mL) with tested strains of bacteria. Among
gram-positive bacteria, complexes were more active towards B. subtilis than S.
aureus and for gram-negative bacteria complexes were more active for P. aer-
uginosa as compared to E. coli, and overall results show that compounds show
more potency for gram-negative bacteria than gram-positive bacteria due to their
distinctive structure and thin cell wall [57, 58].
1 3

Transition metal (II) complexes of hydrazones derived from…
Table 6 Results of in vitro antimicrobial screening for hydrazones (1–4), mononuclear metal complexes
(5–20), and standard drugs (21, 22). (MIC in μmol/mL)
MIC in μmol/mL
C. no. Compounds
Gram-positive bacteria Gram-negative bacteria Fungi
S. aureus B. subtilis E. coli P. aeruginosa C. albicans A. niger
1
2
3
4
5
6
7
8
HL1
HL2
HL3
HL4
0.0942
0.1892
0.0850
0.1885
0.1885
0.1892
0.0425
0.0942
0.0800
0.0399
0.0397
0.0794
0.0804
0.0401
0.0797
0.0795
0.0366
0.0365
0.0091
0.0158
0.0400
0.0797
0.0199
0.0397
0.0188
–
0.1885 0.1885
0.1892 0.0946
0.1700 0.0425
0.1885 0.1885
0.0400 0.0800
0.0797 0.0797
0.0397 0.0795
0.0794 0.0397
0.0804 0.0804
0.0802 0.0401
0.0797 0.0399
0.0397 0.0795
0.0732 0.0366
0.0730 0.0365
0.0182 0.0091
0.0317 0.0158
0.0800 0.0400
0.0797 0.0397
0.0199 0.0158
0.0397 0.0793
0.0188 0.0188
0.1885
0.1892
0.0850
0.1885
0.0800
0.0399
0.0199
0.0794
0.0402
0.0401
0.0199
0.0397
0.0366
0.0365
0.0091
0.0158
0.0400
0.0797
0.0199
0.0397
0.0942
0.1892
0.1700
0.1885
0.0800
0.0797
0.0397
0.0794
0.0804
0.0802
0.0399
0.0795
0.0732
0.0730
0.0091
0.0158
0.0800
0.0797
0.0199
0.0397
[Co(L1)2(H2O)2] 0.0800
[Ni(L1)2(H2O)2] 0.0797
[Cu(L1)2(H2O)2] 0.0795
[Zn(L1)2(H2O)2] 0.0397
[Co(L2)2(H2O)2] 0.0804
[Ni(L2)2(H2O)2] 0.0802
[Cu(L2)2(H2O)2] 0.0797
[Zn(L2)2(H2O)2] 0.0397
[Co(L3)2(H2O)2] 0.0366
[Ni(L3)2(H2O)2] 0.0730
[Cu(L3)2(H2O)2] 0.0182
[Zn(L3)2(H2O)2] 0.0158
[Co(L4)2(H2O)2] 0.0800
[Ni(L4)2(H2O)2] 0.0797
[Cu(L4)2(H2O)2] 0.0199
[Zn(L4)2(H2O)2] 0.0793
9
10
11
12
13
14
15
16
17
18
19
20
21
22
Ciprofoxacin
Fluconazole
0.0188
–
–
–
0.0204
0.0204
0.2000
0.1800
0.1600
0.1400
0.1200
0.1000
0.0800
0.0600
0.0400
0.0200
0.0000
S. aureus
B. subtilis
E. coli
P. aeruginosa
C. albicans
A. niger
Compounds
Fig. 6 Graph showing in vitro antimicrobial activity of hydrazones (1–4) and their transition metal com-
plexes (5–20)
1 3

J. Devi et al.
3. In case of antifungal activity, complexes 7, 11, 15, 16 and 19 were found to show
equally good inhibitory efect than standard fuconazole (MIC=0.204 μmol/mL)
having MIC values in the range 0.0091–0.0399 μmol/mL.
In conclusion, compound (HL3) and its copper and zinc complexes 15 and 16 are
found to more active antimicrobial agent in the entire series of compounds tested.
The antibacterial and antifungal evaluation reveals that the transition metal com-
plexes were observed to be more active than their parent hydrazones against same
species under similar environmental conditions, and this enhancement is explained
by on the basis of overtone’s concept of chelation theory as evidenced from liter-
ature data [59, 60]. The fuctuations in the antimicrobial study rely on the difer-
ences in ribosomes in the microbial cell or the permeability of the cell wall. The
lipid membrane surrounding the cell wall favours the movement of any lipid-soluble
materials, and it is known that lipid solubility is an major factor to control the anti-
microbial activity [61], the nature of metal ion, azomethine linkage in compounds
and chelation phenomena that play an efective role in inhibitory activity [62].
Cytotoxicity
Chemotherapy is the branch of science which deals with both localized and metas-
tasized cancers. The compounds (1–20) were tested for their cytotoxic and growth
inhibitory activities (in vitro) against three human cancer cell lines (A549, Hela,
MCF7) and one normal cell line (L6) by the MTT assay. Doxorubicin, one of the
most active anticancer agents, was taken as a reference standard. The response
parameter was determined as the IC50 values, which response to the concentration
needed for 50% inhibition of cell viability [63, 64]. The correlation between the sur-
viving fraction and drug concentration was plotted to get the survival curve of all
the tested cancer cell lines. The trend followed by hydrazones against cell lines is
HL3 >HL2 >HL4 >HL1, where HL3 hydrazone is most active due to electron releas-
ing methyl group attached to benzene ring. In vitro cytotoxicity of the synthesized
compounds (1–20) is tabulated in Table 7 and bar graph representation in Fig. 7a–d.
The cytotoxic activity data reveal that:
1. Anticancer activity against human alveolar adenocarcinoma epithelial cell line
(A549): Compounds 7 and 15 displayed potent activity, with an IC50 value of
13.28 and 12.81 μg/mL. Other compounds 14 and 16 showed good activity with
IC50 <15 μg/mL, and 3, 8, 9, 11, 12, 13 and 19 gave moderate anticancer activ-
ity, while other compounds in the series do not show any activity. The results are
summarized in Table 3 and are represented in Fig. 7a.
2. The obtained results for human cervical carcinoma cell line (HeLa) demonstrated
that compounds 7, 11 and 15 exhibited very potent activity, with an IC50 value of
10.98, 11.05 and 11.82 μg/mL. Other compounds 13, 14 and 15 showed potency
with IC50 ≤15 μg/mL, whereas 8, 9, 10, 12, 16 and 20 showed moderate antican-
cer activity, while remaining compounds do not show signifcant activity. The
results are presented in Fig. 7b.
1 3